EAGER: Chasing the elusive syntrophic partners in direct interspecies electron transfer

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Anaerobic digestion is widely utilized worlwide to treat waste streams and convert them to biogas such as methane. Biogas fom anerobic digestion typically consists of 50-70% methane and thus needs to be treated to remove impurities including carbon dioxide and water vapor. To eliminate the need for addtional treatment and purification of biogas from existing anaerobic digesters, it is critical to understand and quantify the metabolic pathways of the microbial consortial involved in biogas production. The overarching goal of this high-risk and high-reward EAGER project is to characterize and quantify the metabolic pathways that drive methane production during anaerobic digestion. To advance this goal, the Principal Investigator (PI) of this project proposes to carry out an integrated experimental research program to test the hypothesis that interspecies hydrogen transfer (IHT) and direct interspecies electron transfer (DIET) play an equally important role during biogas production by methanogenesis in electrically conductive environments. The successful completion of this EAGER project could provide new fundamental knowledge that could be leveraged to develop and implement new engineering reactor design and operational strategies to increase the methane content of biogas produced by anaerobic digestors. Further benefits to society will be achieved through student education and training including the mentoring of a doctoral student. Interspecies hydrogen transfer (IHT) and direct interspecies electron transfer (DIET) have been shown to contribute to biogas production by methanogenesis in electrically conductive environments. However, a fundamental understanding and quantification of the relative contributions of IHT and DIET to methanogenesis have remained elusive due to the lack of experimental techniques to directly measure the associated microbial metabolisms and activities. The overarching goal of this project is to address this knowledge gap. To advance this goal, the Principal Investigator (PI) of this project hypothesizes that IHT and DIET play an equally important role during biogas production by methanogenesis in electrically conductive environments. This hypothesis is based on the results of preliminary studies by the PI that identified a novel Geobacter species (Candidatus Geobacter eutrophica) that was abundant in anerobic reactors supplied with conductive granular activated carbon. The PI also found that the Candidatus Geobacter eutrophica bacteria actively expressed genes encoding proteins for both extracellular electron transfer and hydrogen metabolism. To test this new hypothesis, the PI proposes to carry out an integrated experimental research program structured around two specific aims: 1) enrich DIET-capable Geobacter bacteria and elucidate their extracellular electron transfer mechanisms (Specific Aim 1) and 2) enrich DIET-capable methanogens and characterize their extracellular electron uptake mechanisms (Specific Aim 2). To enrich the DIET-capable Geobacter and methanogen bacteria, the PI proposes to use electrochemical stimulation in bioelectrochemical systems with specially designed electrodes. By combining cyclic voltammetry with measurements of biogas production, ion chromatography, resonance Raman microscopy and omics (metagenomics and metatranscriptomics), the PI hopes to unravel the metabolic pathways responsible for DIET in both electron-donating and electron-accepting microbial partners. The successful completion of this project has the potential for transformative impact through the generation of new fundamental knowledge to advance the development of new bioprocesses such as electro-methanogenesis that could convert waste streams to biogas with much higher methane yields than existing anerobic digestion reactors.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.